The objectives of this project are to study the metabolic regulation of hyaluronate in order to investigate further its role in the control of specific aspects of tissue development and remodeling. Major emphasis is centered on the purification and characterization of the enzyme hyaluronidase, as a basis for defining its mechanism of action at the cellular level, on hyaluronate substrate. During year 1 of the grant, I have completed the partial purification and characterization of hyaluronidase from cultured chick embryo fibroblasts (Orkin and Toole, 1980, 255, 1036). Two forms of the enzyme were found. The secreted, medium-associated enzyme is rich in sialic aaid and this confers upon it specific characteristics which distinguish it from the cell-associated form. In particular, the secreted form is a more acidic protein which is more thermally stable at neutral pH than the cell-associated enzyme. During year 2 of the grant, the enzyme will be purified to homogeneity by affinity chromatography. The purified material will undergo compositional analyses and be used to prepare antibody. The enzyme will be localized intracellularly by isolation of specific organelle fractions and/or by use of specific antibodies. The mechanism of action of the enzyme at the cellular level will be studied in enzyme and substrate feeding experiments as well as by the use of agents which destabilize lysosomal function.